Worldwide, more than 400 million individuals become infected with sexually transmitted infections (STI) every year. Since the transmission of STIs occurs through the genital mucosa, vaccines against STIs must be effective in inducing strong immunity at these susceptible mucosal surfaces. A major cellular effector population that provides protection from a variety of intracellular sexually transmitted pathogens is the CD8+ cytotoxic T lymphocyte (CTL). Although the immunological processes involved in the generation and maintenance of systemic memory CTL responses have been studied extensively, the mechanism leading to effective CTL immunity in the cervicovaginal mucosa (CVM) is poorly understood. Several reports indicate that the mere induction of effector CD8+ T cell responses is not sufficient to protect the host from mucosal viral challenge. My laboratory has brought together a variety of approaches and reagents necessary to study inductive, effector and memory CD8+ T cell responses in a physiologically relevant genital STI model. Our research during the first funding period led to the discovery that CTL migration to the CVM requires CD4 T help. Remarkably, we demonstrate in the Preliminary Studies section that even fully helped CTL require CD4 T help in order to enter and remain within the CVM after HSV-2 infection. However, CTL entry into other mucosal organs such as the lung did not require CD4 help, revealing two distinct migration behaviors of CTL - permissive (occurs in the absence of CD4, inflammation or antigen) and restricted (requiring CD4 T cells). We provide new data that demonstrate the critical importance of tissue-resident memory T cells in antiviral protection against genital herpes challenge. Based on these findings, the goal of the proposed study is to examine the anatomical, cellular and molecular mechanisms by which the CD4 T cells enable recruitment and establish effective CD8+ T cell immunity in the female genital mucosa, and to use this knowledge to design vaccine strategies against genital HSV-2 infection. We will accomplish these through 3 distinct but interrelated Aims. In the first Aim, we will determine the homing receptors required for CD4 T cell entry into CVM. In the second Aim, we will determine how local dendritic cells educate naive CD8 T cells to express homing receptor for CVM. In the final Aim, we will utilize the knowledge gained in the previous Aims to design a vaccine strategy that leads to the establishment of local CTL memory pool within the CVM, whereby these cells can recognize and eliminate virally infected cells upon challenge. By providing basic understanding of how CTL gains entry into the genital mucosa, these studies will help to establish critical foundation with which to design immunological interventions and preventative measures against genital herpes and other deleterious STI diseases. Since productive establishment of genital CTL responses is important in defense against many types of sexually transmitted infections, the knowledge gained from the proposed studies will provide critical basic and practical insights for vaccine strategies against STIs in general. PUBLIC HEALTH RELEVANCE: Herpes simplex virus-2 (HSV-2), a causative agent of genital herpes, is a highly prevalent (45 million in the USA alone) sexually transmitted infection. Once acquired, HSV-2 causes a life-long incurable debilitating disease for which no vaccines are currently available. This grant application proposes to examine the mechanism by which cytotoxic T lymphocytes are recruited to the genital mucosa during natural infection and design vaccine strategies based on this knowledge - the understanding obtained through the project will help to establish critical foundation with which to design immunological interventions and preventative measures against genital herpes and other deleterious sexually transmitted diseases.